Cancer-reactive immunity can be identified in tumor-bearing hosts; however, the paucity of immunologic danger signals as well as the presence of immunosuppressive factors/cells within the tumor microenvironment appears to tip the balance in favor of tumor progression. We and others have recently found that the addition of immune activating Ab(s) (e.g. anti-OX40, -CD40, -4-1BB, and -CTLA-4) that stimulate both adaptive and innate immunity can tip the balance in favor of tumor immunity and lead to tumor regression. In particular, our group has focused on the immune stimulating properties of agents targeting the OX40 protein in tumor-bearing hosts, which have shown therapeutic promise in several preclinical mouse cancer models. This application focuses on the changes within T cells and macrophages isolated from the tumor microenvironment (TME) that occur following treatment of tumor-bearing mice with OX40 agonists. We hypothesize that immune-specific changes occur within the TME after OX40 agonist administration that reduce local immune suppression leading to T cell-mediated destruction of tumor cells. The specific aims are as follows; 1) To examine the quantitative and qualitative changes in tumor-reactive CD8 T cells within the tumor microenvironment after OX40 agonist treatment, 2) To understand the role that tumor-associated macrophages (TAMs) play in immune suppression of the TME and how OX40 therapy alleviates this suppression and 3) To test whether reshaping the cytokine milieu in the context of OX40 stimulation can create an immune permissive environment within the tumor. OX40-specific augmentation of the immune system has recently increased in relevance, because we have produced clinical grade anti-OX40 and have treated 20 patients as part the first OX40-specific clinical trial (phase I study). Understanding the changes that occur within the TME following OX40 treatment may ultimately discover new pathways and novel mechanisms, which could be the basis for future clinical trials that combine OX40 therapy with other treatment modalities.